Cao Zhigang, Xu Li, Zhao Shuli, Zhu Xu
Department of Urology, Nanjing First Hospital, Nanjing Medical University, Nanjing, China.
Central Laboratory, Nanjing First Hospital, Nanjing Medical University, Nanjing, China.
Onco Targets Ther. 2019 May 7;12:3429-3439. doi: 10.2147/OTT.S193661. eCollection 2019.
To detect the expression of miR-124 in bladder cancer (BC) cell lines and tissue specimens and to analyze its association with the growth of the BC cells. Quantitative real-time polymerase chain reaction (qPCR) was applied to examine the expression of miR-124 in BC cell lines and tissues. The function of miR-124 in modulating cell proliferation was assessed in BC cells with miRNA-124 overexpression; the cell viability was identified by Cell Count Kit-8; flow cytometry was employed to detect the cell cycle; the expressions of E2F3, cyclin-dependent kinase 4 (CDK4), Ki-67 and vascular endothelial growth factor (VEGF) were tested by qPCR and Western blot; angiogenesis experiment was performed to analysis changes in angiogenesis rate; and bioinformatics prediction and dual luciferase reporter system were employed to identify the target of miR-124. Survival curve data showed that the expression of MicroRNA-124 was positively correlated with survival. MicroRNA-124 expression was significantly decreased in BC cell lines and tissues. Bioinformatics prediction and dual luciferase reporter system verified CDK4 as a direct target of miR-124, which regulated the proliferation of BC cells by directly inhibiting CDK4. BC cells over-expressing miR-124 showed significantly inhibited cell viability, decreased angiogenesis rate, prevented cell proliferation and diminished the expression of E2F3, CDK4, Ki-67 and VEGF. All of these changes were reversed by over-expressing CDK4. MicroRNA-124 suppressed the proliferation of CRC cells by directly targeting CDK4, which provides a target for improving the therapeutic effect of BC.
检测miR-124在膀胱癌（BC）细胞系和组织标本中的表达，并分析其与BC细胞生长的关系。应用定量实时聚合酶链反应（qPCR）检测miR-124在BC细胞系和组织中的表达。在过表达miRNA-124的BC细胞中评估miR-124调节细胞增殖的功能；通过细胞计数试剂盒-8鉴定细胞活力；采用流式细胞术检测细胞周期；通过qPCR和蛋白质免疫印迹法检测E2F3、细胞周期蛋白依赖性激酶4（CDK4）、Ki-67和血管内皮生长因子（VEGF）的表达；进行血管生成实验以分析血管生成率的变化；采用生物信息学预测和双荧光素酶报告系统鉴定miR-124的靶标。生存曲线数据显示，MicroRNA-124的表达与生存率呈正相关。MicroRNA-124在BC细胞系和组织中的表达显著降低。生物信息学预测和双荧光素酶报告系统验证CDK4是miR-124的直接靶标，其通过直接抑制CDK4来调节BC细胞的增殖。过表达miR-124的BC细胞显示出细胞活力显著抑制、血管生成率降低、细胞增殖受到抑制以及E2F3、CDK4、Ki-67和VEGF的表达减少。过表达CDK4可逆转所有这些变化。MicroRNA-124通过直接靶向CDK4抑制CRC细胞的增殖，这为提高BC的治疗效果提供了一个靶点。
